Treating osteoporosis with systemic growth factors and cytokines has not resulted in the expected improvement in bone mass as was initially predicted based on cell culture studies. It is becoming increasing realized that it is the growth factors that are deposited within the bone matrix, possibly as new bone is being formed, that may influence the metabolic activity of subsequent generations of osteoblasts. Thus delivering growth factors to the bone matrix might have potential therapeutic advantages over other routes of administration. Somatic gene therapy in which transplanted osteoblasts are engineered to express a specific growth or cytokine modifying factor is one approach to get production of these factors at the time of new born formation. This project is closely linked with projects 1 and 3 which will determine if transgenic expression of IL-1ra or TNFbp and IGF1 will reduce the bone loss associated with ovariectomy or glucocorticoids. Our project will determine if the same beneficial effect can be obtained by transplantation of osteoprogenitor expressing these factors. However, significant problems which may be unique to bone need serious consideration before a cogent strategy for somatic gene therapy can be formulated. Five fundamental questions will be asked utilizing transgenic donor mice bearing marker genes which can indicate the source and the differentiation state of cells derived from a manipulated stem cell. (1) Do renewing osteoblasts naturally arise from a circulating stem cell, a tissue resident stem cells or both? For stem cell therapy to be successful, engineered cells will need to enter a natural pool of circulating cells to populate bone and be able to synthesize a bony matrix. These studies will be answered with allophanic mice derived from embryo donors bearing different markers ransgenes. We will determine if the two marked populations intermingle or are randomly segregated. If they are segregated, do the growth factors made by one population influence the cells which do not make the factor? (2) Even if stem cells do not normally circulate, can stem cells be made to populate bone and yield differentiated osteoblasta? MSC will be expanded in vitro and used in a heterotopic and intramedullary assay of bone formation and stem cell number. A mouse engineered with TK gene controlled by the Col1A1 promoter will be used to assess the ability of transplanted MSC to engraft bone and participate in bone formation. (3) and (4) Can MSC be manipulated with retroviral or BPV vectors containing either ubiquitous or a Col1A1 promoter and still maintain its ability to home to bone and make bone? This grant is a fusion of three PI's bringing different but interacting disciplines to the complex question of feasibility of MSCs as the cellular ingredient in an overall strategy of somatic gene therapy of bone.